I am involved in a small project to isolate Australian
strains of the protozoan Neospora, which causes abortion and
perinatal mortality in cattle. One component of the project
involves inoculating cell cultures with tissue homogenates
from fresh affected foetuses. Immunohistochemistry and PCR
is also being done by others in the project for detection of
Neospora in fixed tissues.
I have a need to store portions of tissues containing viable
protozoal cysts that can be thawed at a later stage in an
attempt to culture Neospora. For example, we will be
collecting tissues from many sites of an adult cow suspected
to be a carrier of Neospora. Following immunohistochemical
or PCR detection of the organism in specific tissues, it is
envisaged that I could go back to stored viable samples from
the same tissue site and attempt to isolate Neospora in
cell culture. Otherwise the number of cell cultures would
be overwhelming.
So far, I have only had experience freezing viable cultured
cells. What are some suitable methods for preparing tissue
containing protozoa that could be stored in a viable state?
I have access to liquid nitrogen and -80 degrees Celsius
freezer storage space and am assuming that the former would
be necessary, but would appreciate any advice to the
contrary (for example, we store our virus stocks at -80
degrees Celsius). Would it be necessary to add storage
medium to the samples? Should the samples be homogenised
before freezing down and, if so, should they be homogenised
"dry" (without added medium) or in storage medium? What are
appropriate freezing down rates for such samples?
Thanks for your assistance.
Adrian W Philbey
Veterinary Research Officer
Elizabeth Macarthur Agricultural Institute
Private Mail Bag 8
Camden NSW 2570
Australia
Telephone: 61-46-293332
Facsimile: 61-46-293429
email: philbea at agric.nsw.gov.au